This invention relates to genetically engineered strains of bacteria of the genus Streptomyces. As used in this application, the term "Streptomyces bacteria" or "Streptomyces" means any bacterial strain that is a member of the genus Streptomyces as classified in Buchanan et al., The Shorter Bergey's Manual For Determinative Bacteriology (Williams & Wilkins 1982). The invention also relates to vectors for engineering Streptomyces and methods of producing desired compounds with the resulting engineered organisms.
For some time now, the pharmaceutical industry has used strains of Streptomyces to produce desired compounds, particularly antibiotics.
Vectors carrying various genes that are expressed in Streptomyces have been reported. Katz et al. (1983) J. General Microbiol. 129:2703-2714 report cloning a DNA fragment coding for tyrosinase from S. antibioticus DNA into two plasmids. The resulting hybrid plasmids are inserted in a strain of S. lividans. They report that most of the tyrosinase activity of S. antibioticus is secreted; in contrast, most of the activity remains intracellular in S. lividans clones. There is no indication of the mechanism for transport of the activity through the cell wall into the medium.
Thompson et al. (1980) Nature 286:525-527 report cloning the tsr gene from S. azureus which is expressed under its own promoter on various vectors.
Thompson et al. (1983) PNAS (USA) 80:5190-5194 report sequencing the S. fradiae aph (neomycin resistance) gene together with its promoter region. They also report that the aph gene has been incorporated into high copy-number, broad host-range vectors.
Burnett et al. (1984) Abstracts of the Annual Meeting of the American Society of Microbiology H98, p. 107, report cloning a 10-kb fragment from S. lividans containing a transcription start site and two structural genes (beta-galactosidase and Bgl protein). They have determined the sequence of that promoter region as well as of a ribosome-binding site.
Robbins et al. (1981) J. Biol. Chem. 256:10,640-10,644 report cloning the gene coding for the enzyme endo-N-acetylglucosaminidase (endo H) from S. plicatus into E. coli plasmid pBR322. The enzyme is expressed in E. coli and is found in the cytoplasmic, periplasmic, and supernatant fractions.
There have been reports of vectors that include a leader sequence which enables secretion of an expressed protein in other bacterial species. For example, see Gilbert U.S. Pat. Nos. 4,338,397 and 4,411,994 [E. coli]; and Palva et al. (1983) Gene 22:229-235 [Bacillus subtilis].